The action of botulinum neurotoxins (BoNTs) at the neuromuscular junction has been extensively investigated and knowledge gained in this field laid the foundation for the use of BoNTs in human pathologies characterized by excessive muscle contractions. Although much more is known about the
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The action of botulinum neurotoxins (BoNTs) at the neuromuscular junction has been extensively investigated and knowledge gained in this field laid the foundation for the use of BoNTs in human pathologies characterized by excessive muscle contractions. Although much more is known about the action of BoNTs on the peripheral system, growing evidence has demonstrated several effects also at the central level. Pain conditions, with special regard to neuropathic and intractable pain, are some of the pathological states that have been recently treated with BoNTs with beneficial effects. The knowledge of the action and potentiality of BoNTs utilization against pain, with emphasis for its possible use in modulation and alleviation of chronic pain, still represents an outstanding challenge for experimental research. This review highlights recent findings on the effects of BoNTs in animal pain models.
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Although advances in understanding of the pathogenesis of amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA) have suggested attractive treatment strategies, delivery of agents to motor neurons embedded within the spinal cord is problematic. We have designed a strategy based on the
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Although advances in understanding of the pathogenesis of amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA) have suggested attractive treatment strategies, delivery of agents to motor neurons embedded within the spinal cord is problematic. We have designed a strategy based on the specificity of botulinum toxin, to direct entry of viral vectors carrying candidate therapeutic genes into motor neurons. We have engineered and expressed fusion proteins consisting of the binding domain of botulinum toxin type A fused to streptavidin (SAv). This fusion protein will direct biotinylated viral vectors carrying therapeutic genes into motor nerve terminals where they can enter the acidified endosomal compartments, be released and undergo retrograde transport, to deliver the genes to motor neurons. Both ends of the fusion proteins are shown to be functionally intact. The binding domain end binds to mammalian nerve terminals at neuromuscular junctions, ganglioside GT1b (a target of botulinum toxin), and a variety of neuronal cells including primary chick embryo motor neurons, N2A neuroblastoma cells, NG108-15 cells, but not to NG CR72 cells, which lack complex gangliosides. The streptavidin end binds to biotin, and to a biotinylated Alexa 488 fluorescent tag. Further studies are in progress to evaluate the delivery of genes to motor neurons in vivo, by the use of biotinylated viral vectors.
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Spiders are the most successful venomous animals and the most abundant terrestrial predators. Their remarkable success is due in large part to their ingenious exploitation of silk and the evolution of pharmacologically complex venoms that ensure rapid subjugation of prey. Most spider venoms
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Spiders are the most successful venomous animals and the most abundant terrestrial predators. Their remarkable success is due in large part to their ingenious exploitation of silk and the evolution of pharmacologically complex venoms that ensure rapid subjugation of prey. Most spider venoms are dominated by disulfide-rich peptides that typically have high affinity and specificity for particular subtypes of ion channels and receptors. Spider venoms are conservatively predicted to contain more than 10 million bioactive peptides, making them a valuable resource for drug discovery. Here we review the structure and pharmacology of spider-venom peptides that are being used as leads for the development of therapeutics against a wide range of pathophysiological conditions including cardiovascular disorders, chronic pain, inflammation, and erectile dysfunction.
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There is mounting evidence to suggest that environmental factors play a major role in the development of neurodegenerative diseases like ALS (Amyotrophic Lateral Sclerosis). The non-protein amino acid beta-N-methylamino-L-alanine (BMAA) was first associated with the high incidence of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex
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There is mounting evidence to suggest that environmental factors play a major role in the development of neurodegenerative diseases like ALS (Amyotrophic Lateral Sclerosis). The non-protein amino acid beta-N-methylamino-L-alanine (BMAA) was first associated with the high incidence of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia Complex (ALS/PDC) in Guam, and has been implicated as a potential environmental factor in ALS, Alzheimer’s disease, and other neurodegenerative diseases. BMAA has a number of toxic effects on motor neurons including direct agonist action on NMDA and AMPA receptors, induction of oxidative stress, and depletion of glutathione. As a non-protein amino acid, there is also the strong possibility that BMAA could cause intraneuronal protein misfolding, the hallmark of neurodegeneration. While an animal model for BMAA-induced ALS is lacking, there is substantial evidence to support a link between this toxin and ALS. The ramifications of discovering an environmental trigger for ALS are enormous. In this article, we discuss the history, ecology, pharmacology and clinical ramifications of this ubiquitous, cyanobacteria-derived toxin.
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Aggregatibacter actinomycetemcomitans is an oral pathogen and etiologic agent of localized aggressive periodontitis. The bacterium is also a cardiovascular pathogen causing infective endocarditis. A. actinomycetemcomitans produces leukotoxin (LtxA), an important virulence factor that targets white blood cells (WBCs) and plays a role in
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Aggregatibacter actinomycetemcomitans is an oral pathogen and etiologic agent of localized aggressive periodontitis. The bacterium is also a cardiovascular pathogen causing infective endocarditis. A. actinomycetemcomitans produces leukotoxin (LtxA), an important virulence factor that targets white blood cells (WBCs) and plays a role in immune evasion during disease. The functional receptor for LtxA on WBCs is leukocyte function antigen-1 (LFA-1), a b-2 integrin that is modified with N-linked carbohydrates. Interaction between toxin and receptor leads to cell death. We recently discovered that LtxA can also lyse red blood cells (RBCs) and hemolysis may be important for pathogenesis of A. actinomycetemcomitans. In this study, we further investigated how LtxA might recognize and lyse RBCs. We found that, in contrast to a related toxin, E. coli a-hemolysin, LtxA does not recognize glycophorin on RBCs. However, gangliosides were able to completely block LtxA-mediated hemolysis. Furthermore, LtxA did not show a preference for any individual ganglioside. LtxA also bound to ganglioside-rich C6 rat glioma cells, but did not kill them. Interaction between LtxA and C6 cells could be blocked by gangliosides with no apparent specificity. Gangliosides were only partially effective at preventing LtxA-mediated cytotoxicity of WBCs, and the effect was only observed when a high ratio of ganglioside:LtxA was used over a short incubation period. Based on the results presented here, we suggest that because of the similarity between N-linked sugars on LFA-1 and the structures of gangliosides, LtxA may have acquired the ability to lyse RBCs.
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Wheat grains are well known to be invaded by Fusarium spp. under field and storage conditions and contaminated with fumonisins. Therefore, determining Fusarium spp. and fumonisins in wheat grains is of prime importance to develop suitable management strategies and to minimize risk. Eighty-two
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Wheat grains are well known to be invaded by Fusarium spp. under field and storage conditions and contaminated with fumonisins. Therefore, determining Fusarium spp. and fumonisins in wheat grains is of prime importance to develop suitable management strategies and to minimize risk. Eighty-two stored wheat samples produced in Iran were collected from various supermarkets and tested for the presence of Fusarium spp. by agar plate assay and fumonisins by HPLC. A total of 386 Fusarium strains were isolated and identified through morphological characteristics. All these strains belonged to F. culmorum, F. graminearum, F. proliferatum and F.verticillioides. Of the Fusarium species, F. graminearum was the most prevalent species, followed by F. verticillioides, F. proliferatum and then F. culmorum. Natural occurrence of fumonisin B1 (FB1) could be detected in 56 (68.2%) samples ranging from 15–155 μg/kg, fumonisin B2 (FB2) in 35 (42.6%) samples ranging from 12–86 μg/kg and fumonisin B3 (FB3) in 26 (31.7%) samples ranging from 13–64 μg/kg. The highest FB1 levels were detected in samples from Eilam (up to 155 μg/kg) and FB2 and FB3 in samples from Gilan Gharb (up to 86 μg/kg and 64 μg/kg).
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Botulinum neurotoxins are highly effective therapeutic products. Their therapeutic success results from highly specific and potent inhibition of neurotransmitter release with a duration of action measured in months. These same properties, however, make the botulinum neurotoxins the most potent acute lethal toxins known.
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Botulinum neurotoxins are highly effective therapeutic products. Their therapeutic success results from highly specific and potent inhibition of neurotransmitter release with a duration of action measured in months. These same properties, however, make the botulinum neurotoxins the most potent acute lethal toxins known. Their toxicity and restricted target cell activity severely limits their clinical utility. Understanding the structure-function relationship of the neurotoxins has enabled the development of recombinant proteins selectively incorporating specific aspects of their pharmacology. The resulting proteins are not neurotoxins, but a new class of biopharmaceuticals, Targeted Secretion Inhibitors (TSI), suitable for the treatment of a wide range of diseases where secretion plays a major role. TSI proteins inhibit secretion for a prolonged period following a single application, making them particularly suited to the treatment of chronic diseases. A TSI for the treatment of chronic pain is in clinical development.
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Shiga toxin-producing Escherichia coli is a contaminant of food and water that in humans causes a diarrheal prodrome followed by more severe disease of the kidneys and an array of symptoms of the central nervous system. The systemic disease is a complex referred
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Shiga toxin-producing Escherichia coli is a contaminant of food and water that in humans causes a diarrheal prodrome followed by more severe disease of the kidneys and an array of symptoms of the central nervous system. The systemic disease is a complex referred to as diarrhea-associated hemolytic uremic syndrome (D+HUS). D+HUS is characterized by thrombocytopenia, microangiopathic hemolytic anemia, and acute renal failure. This review focuses on the renal aspects of D+HUS. Current knowledge of this renal disease is derived from a combination of human samples, animal models of D+HUS, and interaction of Shiga toxin with isolated renal cell types. Shiga toxin is a multi-subunit protein complex that binds to a glycosphingolipid receptor, Gb3, on select eukaryotic cell types. Location of Gb3 in the kidney is predictive of the sites of action of Shiga toxin. However, the toxin is cytotoxic to some, but not all cell types that express Gb3. It also can cause apoptosis or generate an inflammatory response in some cells. Together, this myriad of results is responsible for D+HUS disease.
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Vectors based on adeno-associated virus (AAV) are promising tools for gene therapy. The production of strongly toxic vectors, for example for cancer-directed gene transfer, is often unfeasible due to uncontrolled expression of toxic genes in vector-producing cells. Using an approach based on transcriptional
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Vectors based on adeno-associated virus (AAV) are promising tools for gene therapy. The production of strongly toxic vectors, for example for cancer-directed gene transfer, is often unfeasible due to uncontrolled expression of toxic genes in vector-producing cells. Using an approach based on transcriptional repression, we have created novel AAV vectors carrying the genes coding for diphtheria toxin A (DTA) and the pro-apoptotic PUMA protein. The DTA vector had a significant toxic effect on a panel of tumor cell lines, and abrogation of protein synthesis could be shown. The PUMA vector had a toxic effect on HeLa and RPMI 8226 cells, and sensitized transduced cells to doxorubicin. To permit targeted gene transfer, we incorporated the DTA gene into a genetically modified AAV-2 capsid previously developed by our group that mediates enhanced transduction of murine breast cancer cells in vitro. This vector had a stronger cytotoxic effect on breast cancer cells than DTA vectors with wildtype AAV capsid or vectors with a random capsid modification. The vector production and application system presented here allows for easy exchange of promotors, transgenes and capsid specificity for certain target cells. It will therefore be of great possible value in a broad range of applications in cytotoxic gene therapy and significantly broadens the spectrum of available tools for AAV-based gene therapy.
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The peanut leaf spot pathogen Passalora arachidicola (Mycosphaerella arachidis) is known to produce dothistromin, a mycotoxin related to aflatoxin. This is a feature shared with the pine needle pathogen Dothistroma septosporum (Mycosphaerella pini). Dothistromin biosynthesis in D. septosporum commences at an unusually early stage of growth in culture compared to most other fungal secondary metabolites, and the biosynthetic genes are arranged in fragmented groups, in contrast to aflatoxin gene clusters. Dothistromin biosynthetic genes were identified and studied in P. arachidicola to determine if the attributes described in D. septosporum are shared by another dothistromin-producing species within the Class Dothideomycetes. It was shown that dothistromin biosynthesis is very similar in the two species with regard to gene sequence and gene synteny. Functional complementation of D. septosporum mutants with P. arachidicola dothistromin genes was also possible. These similarities support a vertical mode of dothistromin gene transmission. P. arachidicola also produced dothistromin at an early growth stage in culture, suggesting that this type of regulation pattern may be relevant to the biological role of dothistromin.
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